The Effects of Model Insoluble Copper Compounds in a Sedimentary Environment on Denitrifying Anaerobic Methane Oxidation (DAMO) Enrichment

The contribution of denitrifying anaerobic methane oxidation (DAMO) as a methane sink across different habitats, especially those affected by anthropogenic activities, remains unclear. Mining and industrial and domestic use of metals/metal-containing compounds can all cause metal contamination in freshwater ecosystems. Precipitation of metal ions often limits their toxicity to local microorganisms, yet microbial activity may also cause the redissolution of various precipitates. In contrast to most other studies that apply soluble metal compounds, this study investigated the responses of enriched DAMO culture to model insoluble copper compounds, malachite and covellite, in simulated sedimentary environments. Copper ≤ 0.22 µm from covellite appeared to cause immediate inhibition in 10 h. Long-term tests (54 days) showed that apparent methane consumption was less impacted by various levels of malachite and covellite than soluble copper. However, the medium-/high-level malachite and covellite caused a 46.6-77.4% decline in denitrification and also induced significant death of the representative DAMO microorganisms. Some enriched species, such as <i>Methylobacter tundripaludum</i>, may have conducted DAMO or they may have oxidized methane aerobically using oxygen released by DAMO bacteria. Quantitative polymerase chain reaction analysis suggests that <i>Candidatus</i> Methanoperedens spp. were less affected by covellite as compared to malachite while <i>Candidatus</i> Methylomirabilis spp. responded similarly to the two compounds. Under the stress induced by copper, DAMO archaea, <i>Planctomycetes</i> spp. or <i>Phenylobacterium</i> spp. synthesized PHA/PHB-like compounds, rendering incomplete methane oxidation. Overall, the findings suggest that while DAMO activity may persist in ecosystems previously exposed to copper pollution, long-term methane abatement capability may be impaired due to a shift of the microbial community or the inhibition of representative DAMO microorganisms.